Technical Challenges for CTC Implementation in Breast Cancer

Abstract

Breast cancer is the most common neoplasm in women worldwide. Tissue biopsy, currently the gold standard to obtain tumor molecular information, is invasive and might be affected by tumor heterogeneity rendering it incapable to portray the complete dynamic picture by the absence of specific genetic changes during the evolution of the disease. In contrast, liquid biopsy can provide unique opportunities for real-time monitoring of disease progression, treatment response and for studying tumor heterogeneity combining the information of DNA that tumors spread in the blood (circulating tumor DNA) with CTCs analysis. In this review, we analyze the technical and biological challenges for isolation and characterization of circulating tumor cells from breast cancer patients. Circulating tumor cell (CTC) enumeration value is included in numerous clinical studies due to the prognostic's role of these cells. Despite this, there are so many questions pending to answer. How to manage lymphocytes background, how to distinguish the CTCs subtypes or how to work with frozen samples, are some of the issues that will discuss in this review. Based on our experience, we try to address these issues and other technical limitations that should be solved to optimize the standardization of protocols, sample extraction procedures, circulating-tumor material isolation (CTCs vs. ctDNA) and the very diverse methodologies employed, aiming to consolidate the use of CTCs in the clinic. Furthermore, we think that new approaches focusing on isolation CTCs in other body fluids such as cerebrospinal or ascitic fluid are necessary to increase the opportunities of circulating tumor cells in the practice clinic as well as to study the promising role of CTC clusters and their prognostic value in metastatic breast cancer.

Keywords: CTC clusters; CTCs; body fluids; breast cancer; liquid biopsy.

Figure 1

Differences between liquid biopsies and conventional tissue biopsies. Image partially created with BioRender.com and modified from Eslami-S et al. [14].

Figure 2

Blood dissemination of different CTCs-subtypes. Primary tumors can shed several CTCs, but only a minim percentage can survive in the bloodstream and only a minority is able to initiate metastatic tumor growth in a distant organ. Mirroring complex heterogeneity of primary tumor cells, phenotypically distinct CTCs-subtypes can co-exist in the circulation. Image created with BioRender.com and modified from Margherita Correnti and Chiara Raggi [40].

Figure 3

Body fluids as a source of tumor-derives molecular information. Schematic representation of different body fluids (other than blood) that can contain tumor-derived molecular information. Specifically cerebrospinal fluid (CSF), pleural effusions, saliva, urine. Image created with BioRender.com, and modified from Siravegna et al. [74].